Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V000415 | Tet-on circRNA Expression Vector | In stock, 1 week for quality controls |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
Low Ampicillin. The vector can efficiently produce a circRNA (circmCherry) from a split EGFP gene after Dox treatment.
- Vector Name:
- Tet-on circRNA Expression Vector
- Antibiotic Resistance:
- Ampicillin
- Length:
- 10054 bp
- Type:
- Mammalian Expression, Lentiviral
- Replication origin:
- ori
- Copy Number:
- High Copy
- Promoter:
- tight TRE
- Cloning Method:
- Restriction Enzyme
- 5' Primer:
- none specified
Tet-on circRNA Expression Vector vector Vector Map
Plasmid Protocol
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.
3. Close the tube and incubate for 10 minutes at room temperature.
4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
5. Store the plasmid at -20 ℃.
6. The concentration of plasmid re-measurement sometimes differs from the nominal value, which may be due to the position of the lyophilized plasmid in the tube, the efficiency of the re-dissolution, the measurement bias, and adsorption on the wall of the tube, therefore, it is recommended to transform and extract the plasmid before using it
General Plasmid Transform Protocol
1. Take one 100μl of the competent cells and thaw it on ice for 10min, add 2μl of plasmid, then ice bath for 30min, then heat-shock it at 42℃ for 60s, do not stir, and then ice bath for 2min.
2. Add 900μl of LB liquid medium without antibiotics, and incubate at 37℃ for 45min (30℃ for 1-1.5 hours) with 180rpm shaking.
3. Centrifuge at 6000rpm for 5min, leave only 100μl of supernatant to resuspend the bacterial precipitate and spread it onto the target plasmid-resistant LB plate.
4. Invert the plate and incubate at 37℃ for 14h, or at 30℃ for 20h.
5. Pick a single colony into LB liquid medium, add the corresponding antibiotics, incubate at 220rpm for 14h, and extract the plasmid according to the experimental needs and the instructions of the plasmid extraction kit.
References
- Zhang Y, Xue W, Li X, et al. The Biogenesis of Nascent Circular RNAs. Cell Rep. 2016;15(3):611-624. doi:10.1016/j.celrep.2016.03.058
Tet-on circRNA Expression Vector vector Sequence
LOCUS V000415 10054 bp DNA circular SYN 13-MAY-2021
DEFINITION Exported.
ACCESSION V000415
VERSION V000415
KEYWORDS Tet-on circRNA Expression Vector
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
.
REFERENCE 1 (bases 1 to 10054)
AUTHORS Zhang Y, Xue W, Li X, Zhang J, Chen S, Zhang JL, Yang L, Chen LL
TITLE The Biogenesis of Nascent Circular RNAs.
JOURNAL Cell Rep. 2016 Apr 19;15(3):611-24. doi:
10.1016/j.celrep.2016.03.058. Epub 2016 Apr 7.
PUBMED 27068474
REFERENCE 2 (bases 1 to 10054)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 10054)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; doi:
"10.1016/j.celrep.2016.03.058"; journalName: "Cell Rep"; date:
"2016-04-19- 19"; volume: "15"; issue: "3"; pages: "611-24"
SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..10054
/mol_type="other DNA"
/organism="synthetic DNA construct"
LTR 1..634
/label="3' LTR"
/note="3' long terminal repeat (LTR) from HIV-1"
misc_feature 681..806
/label="HIV-1 Psi"
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 1303..1536
/label="RRE"
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 1721..1765
/label="gp41 peptide"
/note="antigenic peptide corresponding to amino acids 655
to 669 of the HIV envelope protein gp41 (Lutje Hulsik et
al., 2013)"
CDS 1914..1955
/note="Protein Tat from Human immunodeficiency virus type 1
group M subtype B (isolate WMJ22). Accession#: P12509"
/label="Protein Tat"
misc_feature 2058..2175
/label="cPPT/CTS"
/note="central polypurine tract and central termination
sequence of HIV-1"
promoter 2227..2541
/label="tight TRE promoter"
/note="Tet-responsive promoter PTight, consisting of seven
tet operator sequences followed by the minimal CMV
promoter"
regulatory 2569..2578
/label="Kozak sequence"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
/regulatory_class="other"
primer_bind complement(2620..2641)
/label="EGFP-N"
/note="EGFP, reverse primer"
primer_bind 3942..3961
/label="mCherry-F"
/note="mCherry, forward primer"
misc_feature 4284..4870
/label="IRES2"
/note="internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)"
primer_bind complement(5890..5909)
/label="EXFP-R"
/note="For distinguishing EGFP variants, reverse primer"
primer_bind 6237..6258
/label="EGFP-C"
/note="EGFP, forward primer"
misc_feature 6444..7032
/label="WPRE"
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
LTR 7107..7340
/label="3' LTR (Delta-U3)"
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
primer_bind complement(7459..7477)
/label="pBRforEco"
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 7545..7649
/label="AmpR promoter"
CDS 7650..8507
/label="AmpR"
/note="beta-lactamase"
rep_origin 8681..9269
/label="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 9423..9440
/label="L4440"
/note="L4440 vector, forward primer"
primer_bind 9532..9551
/label="SV40pro-F"
/note="SV40 promoter/origin, forward primer"
protein_bind 9680..9701
/label="CAP binding site"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 9716..9746
/label="lac promoter"
/note="promoter for the E. coli lac operon"
protein_bind 9754..9770
/label="lac operator"
/bound_moiety="lac repressor encoded by lacI"
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
primer_bind 9759..9781
/label="M13/pUC Reverse"
/note="In lacZ gene"
primer_bind 9778..9794
/label="M13 rev"
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 9778..9794
/label="M13 Reverse"
/note="In lacZ gene. Also called M13-rev"
primer_bind 9844..9863
/label="EBV-rev"
/note="SV40 polyA terminator, reverse primer"